Department of Molecular Biology and Biotechnology, Firth Court, Western Bank, University of Sheffield, Sheffield, S10 2TN, UK.
Traffic. 2012 Feb;13(2):317-28. doi: 10.1111/j.1600-0854.2011.01311.x. Epub 2011 Dec 6.
Dynamins are a conserved family of proteins involved in many membrane fusion and fission events. Previously, the dynamin-related protein Vps1 was shown to localize to endocytic sites, and yeast carrying deletions for genes encoding both the BAR domain protein Rvs167 and Vps1 had a more severe endocytic scission defect than either deletion alone. Vps1 and Rvs167 localize to endocytic sites at the onset of invagination and disassemble concomitant with inward vesicle movement. Rvs167-GFP localization is reduced in cells lacking vps1 suggesting that Vps1 influences Rvs167 association with the endocytic complex. Unlike classical dynamins, Vps1 does not have a proline-arginine domain that could interact with SH3 domain-containing proteins. Thus, while Rvs167 has an SH3 domain, it is not clear how an interaction would be mediated. Here, we demonstrate an interaction between Rvs167 SH3 domain and the single type I SH3-binding motif in Vps1. Mutant Vps1 that cannot bind Rvs167 rescues all membrane fusion/fission functions associated with Vps1 except for endocytic function, demonstrating the specificity and mechanistic importance of the interaction. In vitro, an Rvs161/Rvs167 heterodimer can disassemble Vps1 oligomers. Overall, the data support the idea that Vps1 and the amphiphysins function together to mediate scission during endocytosis in yeast.
动力蛋白是一个保守的蛋白质家族，参与许多膜融合和裂变事件。先前，已经表明与 dynamin 相关的蛋白 Vps1 定位于内吞部位，并且携带编码 BAR 结构域蛋白 Rvs167 和 Vps1 的基因缺失的酵母比单独缺失任何一个基因都具有更严重的内吞分裂缺陷。Vps1 和 Rvs167 在凹陷开始时定位于内吞部位，并与向内囊泡运动同时解聚。在缺乏 vps1 的细胞中，Rvs167-GFP 的定位减少，这表明 Vps1 影响 Rvs167 与内吞复合物的关联。与经典 dynamin 不同，Vps1 没有可以与 SH3 结构域蛋白相互作用的脯氨酸-精氨酸结构域。因此，虽然 Rvs167 具有 SH3 结构域，但不清楚如何介导相互作用。在这里，我们证明了 Rvs167 SH3 结构域与 Vps1 中的单个 I 型 SH3 结合基序之间的相互作用。不能结合 Rvs167 的突变 Vps1 可以挽救与 Vps1 相关的所有膜融合/裂变功能，除了内吞功能，证明了相互作用的特异性和机制重要性。在体外，Rvs161/Rvs167 异二聚体可以解聚 Vps1 寡聚物。总体而言，数据支持 Vps1 和 amphiphysin 一起在酵母内吞作用中发挥作用以介导分裂的观点。
